Movement of a karyophilic protein through the nuclear pores of oocytes.

Abstract

Large karyophilic proteins are transported across the nuclear envelope in amphibian oocytes. The specific sites within the envelope thorugh which transport occurs and whether molecular size is a limiting factor in the transport process were investigated. The following experimental procedure was employed. Colloidal gold particles, varying in size from .apprx. 20 to 170 .ANG. in diameter were coated with nucleoplasmin, a 165,000 MW karyophilic protein, which is known to be transported through the envelope. The coated gold paticles were microinjected into the cytoplasm of Xenopus oocytes, and the cells were fixed 15 min and 1 h later. The intracellular localization of the Au was then determined with EM. Nucleoplasm-coated particles readily enter the nucleus. On the basis of the distribution of the particles associated with the envelope, transport evidently occurs through the nuclear pores. The size distributions of the Au particles present in the nucleus and cytoplasm were not significantly different, indicating that the envelope does not discriminate among particles with diameters ranging from 50 to 200 .ANG. (the dimensions including the nucleoplasm coat). Colloidal Au coated with trypsin-digested nucleoplasm (which lacks the polypeptide domain required for transport) or exogenous polyvinylpyrrolidone were largely excluded from the nucleus and showed no evidence of transport.